Zorick T S, Echols H
Division of Biochemistry and Molecular Biology, University of California, Berkeley 94720.
J Bacteriol. 1991 Oct;173(19):6307-10. doi: 10.1128/jb.173.19.6307-6310.1991.
The hflA locus of Escherichia coli specifies a multisubunit protease that selectively degrades the cII transcriptional activator of phage lambda. The regulated turnover of cII is critical for the choice between the lytic and lysogenic pathways of viral development. Previous cell fractionation work has indicated that HflA is associated with the inner membrane fraction. We have sought to demonstrate that the HflA protease is localized in the cell membrane of intact cells. To achieve this goal, we have combined electron microscopy of thin-sectioned E. coli cells with antibody tagging by a colloidal gold label. Using antibody to purified HflA protein, we have found preferential membrane labeling for hflA+ cells but not for hflA mutant cells. We conclude that HflA protease is localized in the cell membrane. The membrane location for HflA protein may serve as a component of a targeting mechanism to limit the action of the regulatory protease to selected cytoplasmic proteins.
大肠杆菌的hflA基因座编码一种多亚基蛋白酶，该蛋白酶可选择性降解噬菌体λ的cII转录激活因子。cII的调控性周转对于病毒发育的裂解途径和溶原途径之间的选择至关重要。先前的细胞分级分离工作表明，HflA与内膜部分相关。我们试图证明HflA蛋白酶定位于完整细胞的细胞膜中。为了实现这一目标，我们将大肠杆菌薄切片细胞的电子显微镜检查与胶体金标记的抗体标记相结合。使用针对纯化的HflA蛋白的抗体，我们发现hflA +细胞有优先的膜标记，而hflA突变细胞则没有。我们得出结论，HflA蛋白酶定位于细胞膜中。HflA蛋白的膜定位可能作为靶向机制的一个组成部分，将调节性蛋白酶的作用限制于选定的细胞质蛋白。
